The catalytic and chemical properties of the general acyl CoA dehydrogenase and electron transfer flavoprotein (ETF) and their flavins are being investigated. The mechanism of oxidation of acyl CoA substrates is studied using rapid kinetic and steady state kinetic techniques, suicide and alternate substrates and electron spin resonance (ESR) methods. These techniques, as well as specific covalent modifications of functional groups in the two flavoproteins, are being employed to investigate the interaction between the two proteins and the electron transfer reaction. These approaches are complemented by determinations of the redox potentials of the flavins in the two flavoproteins. Oxidation of an acyl CoA by the dehydrogenase is apparently a 2 electron transfer. Based on the deuterium isotope effect observed in rapid kinetic studies and experiments with suicide substrates, dehydrogenation involves base-catalyzed abstration of a C-2 proton of the thioester followed by electron (or hydride) transfer to the dehydrogenase flavin. Electron transfer to the ETF is a l electron transfer. The interaction between the dehydrogenase and ETF prior to electron transfer may be electrostatic in character based on experiments with a covalently modified dehydrogenase. The redox potentials of the systems are apparently poised to favor the flow of electrons to the ETF. These investigations have contributed to an understanding of acyl CoA flux through the acyl-CoA dehydrogenase-catalyzed step in mitochondrial fatty acid oxidation. This information is important in understanding fatty acid catabolism in normal individuals as well as those persons with hormonally genetically related defects in fatty acid catabolism.